Biology subjects

Altamirano, F.

Publications and source records attributed to Altamirano, F..

4 recordsLinked to original sources

bioRxiv2026

Polycystin-1 C-Terminus Regulates Protein Synthesis-Related Pathways in Cardiomyocytes

Pathologic cardiac hypertrophy requires increased protein synthesis, but the mechanosensors that link membrane stretch to translational control remain poorly understood. Polycystin-1 (PC1), encoded by PKD1, has been proposed as a cardiac mechanosensor, with its C-terminal tail (PC1-CT) promoting hypertrophy in rodent cardiomyocytes. However, its subcellular localization and downstream signaling remain incompletely defined, especially in human cardiomyocytes. Here, we examined endogenous PC1 C-terminus localization and the effects of adenoviral PC1-CT overexpression in human iPSC-derived ventricular cardiomyocytes (hiPSC-CMs) and adult mouse ventricular myocytes. Immunofluorescence revealed a striking striated pattern for both endogenous PC1 C-terminus (detected with a PC1-CT antibody) and the overexpressed PC1-CT fragment. In hiPSC-CMs, the PC1 C-terminus localized between the -actinin bands. In contrast, in adult cardiomyocytes, the overexpressed protein colocalized with -actinin and desmin, suggesting that PC1-CT sarcomeric distribution depends on cardiomyocyte maturation. We performed RNA-seq to assess transcriptional responses downstream of PC1-CT overexpression in hiPSC-CMs relative to LacZ controls. Gene Set Enrichment Analysis (GSEA) revealed enrichment of gene sets related to ribosome biogenesis, RNA processing, and protein synthesis, while classical hypertrophic markers remained unchanged. Pathway analysis suggested increased PI3K activity. PC1-CT overexpression increased phosphorylation of Akt, ERK, S6K1, and ribosomal protein S6 without altering 4EBP1 phosphorylation, suggesting preferential activation of the mTOR-S6K1-S6 branch. Pharmacological studies showed that pan-PI3K inhibition abolished S6 phosphorylation, whereas MEK blockade did not affect it; pertussis toxin and PI3K{gamma}-selective inhibitors also did not affect S6, suggesting a Gi/o-independent PI3K/Akt signaling driving mTOR-S6K1-S6 activation. Collectively, these data identify a sarcomere-associated pool of PC1-CT that engages PI3K-Akt-mTOR-S6K1-S6 signaling to enhance transcriptional programs related to ribosome biogenesis and protein synthesis, without activating a canonical hypertrophic gene program. These findings reveal a mechanistic link between PC1-CT and cardiomyocyte growth.

physiology↗

bioRxiv2026

Human-engineered heart tissues recapitulate tissue-scale mechanisms underlying ventricular tachycardia

Human iPSC-derived engineered heart tissues (EHTs) and cardiac organoids are increasingly used to model cardiac physiology and drug responses, yet it remains unclear whether they can reproduce tissue-scale mechanisms underlying ventricular arrhythmias. Advanced electrophysiological characterization of EHTs has been limited by the lack of a recording framework compatible with small, perfused preparations, despite the availability of mapping hardware. We present a reproducible workflow that couples milliPillar-based EHT fabrication with high-resolution (22 {micro}m spatial, 1 ms temporal) dual channel optical mapping using RH-237 for voltage and Rhod-2 AM for Ca{superscript 2}. Baseline electrophysiological measurements align with published data from human and animal cardiac tissues, showing rate-dependent restitution of action potential and Ca{superscript 2} transient duration (APD and CaD), conduction slowing at higher pacing rates, and physiologic AP-Ca{superscript 2} activation latency. Selective hERG blockade with E-4031 prolongs APD, confirming pharmacological sensitivity. To interrogate mechanisms underlying ventricular tachycardia (VT), we utilized an established proarrhythmic perturbation widely validated in animal models of acquired long-QT syndrome, combining hERG inhibition with hypokalemia and hypomagnesemia; electrolyte disturbances commonly encountered in clinical settings. Treated EHTs (VT group) displayed tachyarrhythmic contractile bursts with marked beat-to-beat instability, whereas controls responded synchronously to field stimulation. Beat-resolved optical mapping revealed progressive diastolic interval shortening, APD dispersion with transient localized long-short APD zones, and regional depression of excitability that together formed spatial conduction barriers precipitating wavebreak and reentry. Early afterdepolarizations contributed to triggered activity and created localized repolarization barriers at long-short APD zones, leading to rotor formation. Phase singularity tracking identified short-lived rotors localized predominantly in the heads of VT EHTs and absent in controls. A minority of tissues exhibited multiple simultaneous rotors and wavelets generating chaotic-like activation. Although tissue acceleration promoted rotor formation, these events were brief, likely due to the spatial limitations of EHTs, and treated tissues more closely resembled VT than Torsades de Pointes or sustained fibrillation. Our comprehensive studies demonstrate that human iPSC-derived EHTs can recapitulate the tissue-scale VT mechanisms associated with acquired long QT syndrome - APD dispersion with long-short APD zones, triggered activity, conduction block, wavebreak and reentry - which had previously been assessed only in intact hearts. The presented platform thus provides a scalable, non-animal system for mechanistic arrhythmia research.

biophysics↗

bioRxiv2024

Polycystin-1 loss of function increases susceptibility to atrial fibrillation through impaired DNA damage response

BackgroundThe increasing prevalence of atrial fibrillation (AF) and chronic kidney diseases highlights the need for a deeper comprehension of the molecular mechanisms linking them. Mutations in PKD1, the gene encoding Polycystin-1 (PKD1 or PC1), account for 85% of autosomal dominant polycystic kidney disease (ADPKD) cases. This disease often includes cardiac complications such as AF. In cardiomyocytes, PC1 deletion reduces hypertrophic response to pressure overload but promotes baseline ventricular dysfunction, while deletion in fibroblasts ameliorates post-myocardial infarction fibrosis. Despite its known cardiac impact, the role of PC1 in atrial cardiomyocytes and arrhythmias is less understood. Here, we sought to investigate the role of PC1 in AF. MethodsWe used intracardiac programmed stimulation and optical mapping to evaluate AF inducibility in two mouse models, Pkd1 R3277C, which recapitulates human ADPKD progression, and cardiomyocyte-specific Pkd1 deletion, and their respective controls. Isolated adult mouse atrial cardiomyocytes, human iPSC-derived atrial cardiomyocytes (hiPSC-aCM), and HL-1 cells served as in vitro cellular models. Molecular mechanisms were evaluated using optical mapping and molecular and biochemical approaches. ResultsLoss-of-function PC1 mutations significantly increased AF susceptibility in vivo and facilitated local reentry in ex vivo left atrial appendages. Comprehensive in vitro experiments supported a direct effect of PC1 in atrial cardiomyocytes. PC1-deficient monolayers exhibited increased arrhythmic events, escalating into reentrant spiral waves post-tachypacing. Transcriptomics analysis revealed PC1-dependent regulation of DNA repair, with PC1 deficiency leading to increased DNA damage under stress. PARP1 inhibitors or nicotinamide riboside, which counteract DNA damage-related metabolic consequences, reduced in vitro arrhythmias PC1-deficient monolayers. Overexpression of the C-terminus of PC1 had the opposite effects in DNA repair genes, suggesting its regulatory effects in atrial cardiomyocytes through retinoblastoma/E2F. Analyses of human atrial tissue from non-ADPKD AF patients showed reduced levels of mature PC1, suggesting a broader relevance of impaired PC1 in AF. ConclusionsImpaired PC1 increases in vivo AF inducibility under programmed electrical stimulation and promotes in vitro arrhythmias in hiPSC-aCM and HL-1 cells. Our findings indicate that PC1 protects against DNA damage to reduce AF susceptibility.

pathology↗

bioRxiv2023

Histone demethylase KDM5 regulates cardiomyocyte maturation by promoting fatty acid oxidation, oxidative phosphorylation, and myofibrillar organization

RationaleHuman pluripotent stem cell-derived CMs (iPSC-CMs) are a valuable tool for disease modeling, cell therapy and to reconstruct the CM maturation process and identify, characterize factors that regulate maturation. The transition from immature fetal to adult CM entails coordinated regulation of the mature gene programming, which is characterized by the induction of myofilament and OXPHOS gene expression among others. Recent studies in Drosophila, C. elegans, and C2C12 myoblast cell lines have implicated the histone H3K4me3 demethylase KDM5 and its homologs, as a potential regulator of developmental gene program and mitochondrial function. We speculated that KDM5 may potentiate the maturation of iPSC-CMs by targeting a conserved epigenetic program that encompass mitochondrial OXPHOS and other CM specific maturation genes. ObjectivesThe purpose of this study is to determine the role of KDM5 in iPSC-CM maturation. Methods and ResultsImmunoblot analysis revealed that KDM5A, B, and C expression was progressively downregulated in postnatal cardiomyocytes and absent in adult hearts and CMs. Additionally, KDM5 proteins were found to be persistently expressed in iPSC-CMs up to 60 days after the onset of myogenic differentiation, consistent with the immaturity of these cells. Inhibition of KDM5 by KDM5-C70 -a pan-KDM5 inhibitor-resulted in differential regulation of 2,372 genes including upregulation of Fatty acid oxidation (FAO), OXPHOS, and myogenic gene programs in iPSC-CMs. Likewise, genome-wide profiling of H3K4me3 binding sites by the CUT&RUN assay revealed enriched H3K4me3 peaks at the promoter regions of FAO, OXPHOS, and sarcomere genes. Consistent with the chromatin and gene expression data, KDM5 inhibition led to increased expression of multiple sarcomere proteins, enhanced myofibrillar organization and improved calcium handling. Furthermore, inhibition of KDM5 increased H3K4me3 deposits at the promoter region of the ESRRA gene, which is known to regulate OXPHOS and cardiomyocyte maturation, and resulted in its increased RNA and protein levels. Finally, KDM5 inhibition increased baseline, peak, and spare oxygen consumption rates in iPSC-CMs. ConclusionsKDM5 regulates the maturation of iPSC-CMs by epigenetically regulating the expression of ESRRA, OXPHOS, FAO, and sarcomere genes and enhancing myofibril organization and mitochondrial function.

genomics↗